Study And Application Of Cutinase Produced By

Eucommia ulmoides Oliv is a kind of special Chinese herbal medicine and important industrial raw materials of plant, it was found that a large number of medicinal ingredients contained in eucommia bark and the leaf which can cure many diseases. Therefore it is very active in the field of natural medicine research and has very broad comprehensive utilization.With the development of enzyme preparation, more and more researchers are agreed with the method of extracting natural ingredients in Eucommiae bark by enzyme engineering. The pathogenicity of plants found that its cutinase can damage the cuticle of the host plants. So our research group have targeted screening a kind of strain which could effective degrade the cuticle of Eucommia ulmoides leaves, the strain identified as Rhodotorula mucilaginosa.After researching on the growth characteristics of Rhodotorula mucilaginosa, it was induced ferment produce cutinase. According to the literature reports, the commonly used inducer is plants’ cutin, the manufactur of cutin was very complex, and the large-scale application of cutinase was restricted.Therefore,the work looking for new inducer that can replace the cutin is imperative.After many tests we found the effect of using linseed oil to do the inducer is good, and the linseed oil is cheap and easy to get, this new discovery can reduce the production cost and work greatly, improve the production efficiency. In order to apply the method to the production practice, this paper has done the following work:1. Compared the induction effect of olive oil, rapeseed oil, linseed oil and cutin, screening suitable inducers to produced cutinase efficient. The contrast found, cutinase activity is higher than other inducers when we choose linseed oil for inducer, it is 1.5 times as without inducer. And we make a detail study on the fermentation conditions of induction to confirm the process parameters.2. The study on enzymatic properties of Rhodotorula mucilaginosa cutinase. Mainly includes the optimum reaction temperature, thermal stability, optimum pH, pH stability, as well as the influence of some metal ions on cutinase activity.3. The enzyme combined extraction. Combined extraction of natural components in Eucommia ulmoides leaves with cutinase, protease, pectinase and cellulase. After removed the cuticle, pectin, cellulose, the enzymatic liquid collection step by step. Then filtering, concentrating, alcohol extraction, and finally load to β-cyclodextrin,made Eucommia ulmoides powder. HPLC was used in analysis and determination of the content of chlorogenic acid in Eucommia ulmoides powder, and confirmed the detection method.